Anionic Polyelectrolyte Hydrogels: Influence on Antibodies Production and Enzyme Activity

Abstract

Recombinant technologies are capable to produce specific molecules of DNA or protein that possess antigenic properties and are safe. However, individual antigen molecules are low-immunogenic, and therefore require conjugation with a compound possessing stronger adjuvant properties [3, 10]. Salts of aluminum, aqueous emulsions of squalene, viruses, virus-like nanoparticles, cationic liposomes, and others are used as adjuvants. A strong immune response in mice has been achieved with complete Freund’s adjuvant, which includes lanolin, vaseline oil and killed mycobacterium tuberculosis [7]. However, it has got negative because of granulomas formation at the injection sites [6] and not suitable for the preparation of vaccines. The aluminum oxide hydrate and aluminum phosphate [2] only are currently authorized for use in most countries as adjuvants. Despite the fact that aluminum compounds are considered as safe, the occurrence of abscesses, eosinophils, granules and allergic manifestations at their application has been observed [1]. Therefore, the problem of expanding the assortment of such drugs continues to be topical and important. The purpose of this study is to compare immunological effect of the created polymers using antigen – BSA model and to investigate their effect on the activity of enzymes of antioxidant defense, as well as ALT and AST. Materials and methods. The PHG MG-4 and MG-8 have been synthesized via the dispersion polymerization of a monomer mixture in heptane (LobaChemie, India), azoisobutyronitrile (AIBN, Merck, Germany) has been used as initiator (5 % per monomers). Glycidyl methacrylate (GMA), butyl acrylate (BA), acrylic acid (AA), and triethylene glycol dimethacrylate (TGMDMA) have been used to obtain microsized PHG. Polymerization has been carried out in the flat bottom dilatometers or reactors at stirring for six hours at 70 ± 0.2 °C pre-filled by argon. The kinetic of the reaction has been studied using dilatometric and gravimetric techniques. Polymer has been separated and washed to remove not reacted monomers. As a result of the polymerization a cross-linked PHG has been received (Fig. 1). The content of the carboxyl groups has been determined by reverse acid-base titration followed by centrifugation and the selection of the liquid phase for analysis, the content of epoxy groups by the reverse titration of the residues of chloric acid in 0.1 N NaOH. TEM images of PHG microparticles have been recorded on JEM-200A electron microscope at accelerating voltage of 200 kV. The hydrodynamic diameter and Z-potential of the PHG particles have been measured by dynamic light scattering on Zetasizer Nano (Malvern, UK) device using noninvasive inverse scattering technology at 25 °C. The concentration of samples has been 0.4 mg/ml. The in vivo study of action of PHG particles has been conducted on mice in accordance with the European Convention for the Protection of Vertebrate Animals (Strasbourg, 1986). Mice of 5 months of age have been divided into 4 groups (2 controls and 2 experimental), n = 5. Animals from the first control group have been injected subcutaneously of 40 μl of 0.9 % isotonic NaCl solution: and the second control group 100 mg/ml BSA (AppliChem GmbH,